PCT/US2007/012799, published as International Patent Application WO/2007/143006, describes the use of OABs having low critical micelle concentrations (predominantly less than about 100 ppm) in an aqueous environment of Minimal Essential Media (MEM) that (i) can lower the surface tension in the aqueous MEM environment to less than about 50 dynes/cm2 and (ii) have the ability to increase metabolic soluble proteins. Creams, lotions, gels and serums comprising specific OABs, each having different properties, are disclosed. Among the properties of the OABs disclosed in WO/2007/143006 are the ability to increase the synthesis of skin matrix proteins (e.g., elastin, fibronectin, collagen) and/or increase cell turnover rates while not causing a concomitant increase in the synthesis of enzymes that degrade these proteins (e.g., matrix metalloproteinases). Additionally, several of the disclosed OABs are taught (i) not to cause an increase in inflammatory proteins (notably interleukin 6 and interleukin 8) and/or (ii) to have broad spectrum antimicrobial activity (i.e., the ability to inhibit the growth or kill a variety of microorganisms).
As discussed in WO/2007/143006, the use of amino acid sequences, both acylated and non-acylated, in topically-applied prescription and non-medicated (i.e., cosmeceutical products skin care products is known in the art. Some such sequences are commercially-available as acylated moieties (e.g., acetyl, myristoyl, palmitoyl). In general, acylation is a well-known technique to those of skill in the art for enhancing penetration of a water-loving or hydrophilic ingredient into the skin. The surface of normal skin is highly hydrophobic preventing significant penetration by hydrophilic substances. However, the properties of an acylated amino acid sequence can vary greatly in terms of toxicity, which, in turn, affects its ultimate usefulness. Surprisingly and unexpectedly, many of the oligomeric acylated biosurfactants of the present invention have comparatively low toxicity to mammalian cells (on the order of LD50>200) while at the same time maintaining a relatively high degree of toxicity for prokaryotic life forms.
Unlike prior art acylated amino acid sequences used in topically-applied products, OABs of the present invention have the ability to increase the synthesis of skin Essential Media (MEM) that (i) can lower the surface tension in the aqueous MEM environment to less than about 50 dynes/cm2 and (ii) have the ability to increase metabolic soluble proteins. Creams, lotions, gels and serums comprising specific OABs, each having different properties, are disclosed. Among the properties of the OABs disclosed in WO/2007/143006 are the ability to increase the synthesis of skin matrix proteins (e.g., elastin, fibronectin, collagen) and/or increase cell turnover rates while not causing a concomitant increase in the synthesis of enzymes that degrade these proteins (e.g., matrix metalloproteinases). Additionally, surprisingly and significantly, OABs of the present invention do not cause an increase in inflammatory proteins, notably interleukin 6 and interleukin 8. This combination of properties makes these compounds uniquely suited to skin care applications.
The ability of OABs of the present invention to effectively wet surfaces at low CMCs confers another surprising and unexpected property—broad spectrum antimicrobial activity. Oligomeric biosurfactants of the present invention have the ability to inhibit the growth or kill a variety of microorganisms, including Escherichia coli (E. coli), Pseudomonas aeruginosa (P. aeruginosa), Pseudomonas cepacia (P. cepacia), Staphylococcus aureus (S. aureus), including methicillin resistant S. aureus (MRSA), including (ATCC BAA-41; ATCC BAA-44; ATCC-BAA-39), Staphylococcus epidermidis (S. epidermidis), and Candida albicans (C. albicans).
The International Search Report for WO/2007/143006 identified five prior art references, four patent references (i) WO 2004/099237 (“Ziegler”); (ii) US Patent Application Publication No. 2004/0229808 (“Owen”); (iii) US Patent Application Publication No. 2005/0142081 (“Breton”) and (iv) US Patent Application Publication No. 2002/0111309 (“Castillo”) and one non-patent article Brown, “Biosurfactants for cosmetic applications” Intl. J. Cosmet. Sci. Vol. 13, No. 2, pp. 61-64 (“Brown”). (Granted US patents and published US patent applications referenced herein are, to the extent pertinent, incorporated by reference.)
The Ziegler WO application teaches the use of specific acylated tripeptide sequences, including Lys-Val-Lys (KVK), Lys-Thr-Lys (KTK), Lys-Val-Arg (KVR), Lys-Leu-Lys (KLK), Lys-Ile-Lys (KIK), Lys-Ala-Lys (KAK), Lys-Ser-Lys (KSK), and Arg-Val-Arg (RVR).
The Breton US application teaches compositions comprising an N-acylaminoamide elastase inhibitor and at least one metalloproteinase inhibitor including adapalene or analogous peptides.
The Owen WO application is now U.S. Pat. No. 7,354,903 and is a continuation of U.S. patent application Ser. No. 09/820,053 filed Mar. 28, 2001, now U.S. Pat. No. 6,875,744. As discussed at Paragraph [0087] of WO/2007/143006, OAGs of the present invention are different from the “FLAK” peptides (i.e., those containing Phenylalanine, Leucine, Alanine, and Lysine residues) described in U.S. Pat. No. 6,875,744 which do not auto-aggregate in solution.
The Castillo US application discloses a method for treating diseases associated with beta-amyloid protein formation, deposition or accumulation, including Alzheimer's disease and Down's syndrome, with one of ten laminin-derived non-acylated polypeptides. Intravenous application is taught to be preferred; however, topical, intraarterial, intraperitoneal, oral, intralymphatic, intramuscular and intralumbar administration are also disclosed. The ISR specifically notes the polypeptides of SEQ ID NO 5 which is comprised of 3,075 amino acid residues. Of the remaining sequences, SEQ ID NO 1 is the shortest—containing 11 amino acids. The next shortest of the remaining disclosed sequences contains more than one hundred amino acid residues. OAGs of the present invention are from three to nine amino acids in length.
The Brown article discloses biosurfactants produced by microorganisms, including species of Bacillus, Penicillium and Cornybacterium. The OAGs of the present invention are short sequences of amino acids, from three to nine residues in length, that are made synthetically.
International Patent Application Publication WO/2007/093839 discloses polypeptides according to the formula: A-(Xaa)n-Lys-X-Lys-B. A is defined as —NH2, NH3+, or NH-D, where D is an acyl group, either biotin or alkyl C2-C22. B is defined as H, O_OR1, or NR2R3, where R1, R2 and R3 are independently either H or an alkyl chain (C1-C24). (Xaa)n is an amino acid chain including any natural or synthetic amino acid, but excluding arginine and lysine, where n is an integer between 0 and 3. X is defined as a chain of two amino acids (Xaa1Xaa2), which may be the same or different, excluding arginine, lysine and excluding Xaa1Xaa2=Thr-Thr, Gly-His and Glu-His, or X is a spacer selected among beta-alanyl (β-Ala), 5-amino-valeroyl (“Ava”), 4-amino-cyclohexanoyl, 4-amino-butyroyl (“Abu”), 6-amino-caproyl (“Aca”) and derivatives thereof. More particularly, WO/2007/093839 discloses acylated tetrapeptides according to the formula A-Lys-Xaa1Xaa2-Lys-B, where neither Xaa nor Xaa2 are Arg or Lys. Also excluded from this formula are the tetrapeptide sequences: Lys-Thr-Thr-Lys; Lys-Gly-His-Lys; and Lys-Glu-His-Lys. The following tetrapeptide sequences acylated with a palmitoyl (“Pal”) or elaidoyl (“Ela”) moiety are specifically disclosed: Pal-Lys-Thr-Phe-Lys; Ela-Lys-Thr-Phe-Lys; Ela-Lys-Thr-Ala-Lys; Pal-Lys-Ava-Lys; Ela-Lys-Ala-Tyr-Lys; Ela-Lys-Phe-Tyr-Lys; Pal-Lys-R-Ala-Lys; Pal-Lys-Abu-Lys; and Pal-Lys-Aca-Lys. The International Search Report issued for WO/2007/093839 cites two prior art references, both of which teach the tetrapeptide sequence Lys-Thr-Phe-Lys (“KTFK”).
One aspect of the present invention is directed to hair care compositions comprising OABs and use of OABs in topically-applied prescription and cosmeceutical products to inhibit or reduce hair loss, to stimulate hair growth, and/or to create the appearance of a healthier, shinier, and/or fuller hair shaft and/or reduce the appearance of thinning hair. This application of OABs of the present invention is not claimed or suggested in WO/2007/143006.
Certain OABs disclosed in WO/2007/143006 having at least two charged amino acid residues have now been found to increase the expression of genes associated with cell proliferation and differentiation—KRT1, KRT14, FGF2, GRN and DEFB1. Without wishing to be bound by a theory, Applicants believe that this surprising and unexpected property contributes, in part, to the ability of OABs of the present invention to help to inhibit or reduce hair loss, to stimulate hair growth, and/or to create the appearance of a healthier, shinier, and/or fuller hair shaft and/or reduce the appearance of thinning hair.
The scientific and patent literature has reported that topical use of peptides stimulates hair growth. A subsection of the Background of Invention in WO/2007/143006 is entitled “Prior Art Amino Acid Sequences Used in Skin Care Products.” Among the prior art references discussed is US Patent Application Publication No. 2006/0067905 which describes a method for treating hair loss by administering oleanolic acid, apigenin and Biotinyl-Gly-His-Lys. Biotinyl tripeptide is formed by grafting vitamin H (biotin) on the tripeptide Gly-His-Lys. As explained in WO/2007/143006, the biotinyl moiety on this compound does not confer sufficient hydrophobicity to produce biosurfactant properties.
International Patent Appln PCT/JP2005/004033 (published as WO/2005/082395) teaches the use of the hexapeptide RPLKPW (Arg-Pro-Leu-Lys-Pro-Trp) to promote hair growth and inhibit hair loss.
International Patent Application PCT/KR2006/004352 (published as WO/2007/049905) teaches the use of a thymosin beta-4 mimicking octapeptide KLKLTETG (Lys-Leu-Lys-Leu-Thr-Glu-Thr-Gln) for promoting hair growth.
U.S. Pat. No. 5,538,945 assigned to Procyte Corp. teaches the topical administration of copper-peptide complexes to stimulate hair growth, particularly in persons having androgenetic alopecia, alopecia greata, female pattern baldness and secondary alopecia. The disclosed copper-peptide complexes conform to the formula [R1—R2—R3]: copper(II), wherein R1 is an amino acid or amino acid derivative; R2 is histidine or arginine; and R3 is at least one amino acid or amino acid derivative joined to R2 by a peptide bond, with the proviso that R1 is not glycyl, alanyl, seryl or valyl when R2 is histidyl and R3 is lysine, lysyl-prolyl-valyl-phenylalanyl-valine, lysyl-valyl-phenylalanylvaline, lysyl-tryptophan, or lysyl-(glycyl)1-2-tryptophan, and with the further proviso that R1 is not lysyl when R2 is histidyl and R3 is glycine, glycyl-prolyl-valyl-phenylalanyl-valine, glycyl-valyl-phenylalanyl-valine, glycyltryptophan, or glycyl-(glycyl)1-2-tryptophan.
International Application PCT/US2003/035558 (published as WO/2004/043415) teaches the topical application of the copper-peptide complexes described in Paragraph [0008] in combination with Minoxidil (6-amino-1,2-dihydro-1-hydroxy-2-imino-4-piperidinopyrimidine).
The hexapeptide Arg-Pro-Leu-Lys-Pro-Trp (RPLKPW) has been reported to stimulate hair growth in shaved mice after feeding at a dose of 1 mg/kg for 2 weeks. Y. Masaaki, et al. “RPLKPW, a Designed Hypotensive Peptide Relaxes Artery and Stimulates Hair Growth via Angiotensin AT2 Receptor” J. Pept. Sci., Vol. 2004, pp. 99-102.
The present invention is directed to OABs not previously disclosed in WO/2007/143006 (SEQ ID NO 1-SEQ ID NO 96) as well as new uses of OABs disclosed in that application (SEQ ID NO 97-SEQ ID NO 129). These OABs are setout in the sequence listing in Paragraph [0019] and have a critical micelle concentration of less than about 200 ppm, preferably less than about 50 ppm) in an aqueous environment of Minimal Essential Media (“MEM”) Solution (as defined below) that reduce the surface tension in the aqueous environment to less than about 50 dynes/cm2. More particularly, the OABs consist essentially of (i) an 8- to 22-membered carbon chain, branched or unbranched, saturated or unsaturated, preferably myristoylated (abbreviated “Myr”) or palmitoylated (abbreviated “Pal”); (ii) three to nine amino acid residues, at least one, preferably at least two of which is/are charged; and (iii) an acid C-terminus or an amide C-terminus.
As used in the present application, by charged amino acid is meant lysine, arginine, aspartic acid and glutamic acid.
Surprisingly, OABs of the present invention have been found to have an ability to increase metabolic soluble proteins by at least about 20% while exhibiting comparatively low toxicity for mammalian cells—preferably, having an LD50 of greater 200 ppm in 37 year-old female fibroblast cells.
Also, surprisingly, many OABs of the present invention have the ability to increase synthesis and/or slow degradation of skin matrix proteins.
The following sequence listing forms part of the specification and is included to further illustrate certain aspects of the present invention. The invention maybe better understood by reference to one or more of these sequences in combination with the detailed description of the invention presented below.
P #SequenceSequence IDP257Myr-GAR-acid P322Myr-AHR-amide P342Myr-RRR-amide P348Myr-KKK-acid P388Myr-DAD-acid P291Myr-KKAL-amideSEQ ID NO 1 P362Myr-KKAL-acidSEQ ID NO 2 P363Myr-LAKK-acidSEQ ID NO 3 P371Myr-IAKK-amideSEQ ID NO 4 P372Myr-LSKK-amideSEQ ID NO 5 P389Myr-DDAD-acidSEQ ID NO 6 P398Pal-AAKR-amideSEQ ID NO 7 P399Myr-GGKR-amideSEQ ID NO 8 P400Myr-AAKR-amideSEQ ID NO 9 P405Myr-KGL-[K]D-amideSEQ ID NO 10 P190Myr-KTTKS-amideSEQ ID NO 11 P228Myr-TKTTK-amideSEQ ID NO 12 P263Pal-KKGEM-acidSEQ ID NO 13 P266Myr-GRKGD-acidSEQ ID NO 14 P293Myr-KKGEM-acidSEQ ID NO 15 P328Dod-KKALK-amideSEQ ID NO 16 P349Myr-KKALK-acidSEQ ID NO 17 P351Myr-KLAKK-acidSEQ ID NO 18 P361Myr-K-[K]D-A-[L]D-amideSEQ ID NO 19 P365Myr-KKLA-[K]D-amideSEQ ID NO 20 P374Myr-KKGLK-amideSEQ ID NO 21 P382Myr-GRGDS-amideSEQ ID NO 22 P387Oct-GRGDS-amideSEQ ID NO 23 P390Myr-KKG1-[K]D-amideSEQ ID NO 24 P394Myr-KKGL-[K]D-amideSEQ ID NO 25 P395Myr-KKSL-[K]D-amideSEQ ID NO 26 P401Myr-KIAK-[K]D-amideSEQ ID NO 27 P414Myr-KLAK-[K]D-amideSEQ ID NO 28 P418Myr-IIIKK-amideSEQ ID NO 29 P252Myr-KLAKKA-acidSEQ ID NO 30 P256Myr-AKKLAK-acidSEQ ID NO 31 P286Myr-d-[LKKALK]-acidSEQ ID NO 32 P296Myr-LKKALK-amideSEQ ID NO 33 P364Myr-PKKALK-amideSEQ ID NO 34 P396Myr-KLAKK-[L]D-amideSEQ ID NO 35 P402Myr-AKKGL-[K]D-amideSEQ ID NO 36 P404Myr-GKKAL-[K]D-amideSEQ ID NO 37 P406Myr-LKKAL-[K]D-acidSEQ ID NO 38 P408Myr-SKKAL-[K]D-amideSEQ ID NO 39 P411Myr-DDSSKK-amideSEQ ID NO 40 P412Myr-DDLAKK-amideSEQ ID NO 41 P417Myr-IIIIIK-amideSEQ ID NO 42 P258Myr-AHR-acid P304Pal-GRK-amide P321Myr-KHR-amide P323Pal-AHR-amide P340Myr-KKK-amide P341Pal-KKK-amide P413Myr-SDD-acid P410Myr-DSDD-acidSEQ ID NO 43 P352Myr-d-[KKALK]-amideSEQ ID NO 44 P358Myr-[K]D-KALK-amideSEQ ID NO 45 P378Myr-KKGIK-amideSEQ ID NO 46 P385Myr-KIAKK-amideSEQ ID NO 47 P409Myr-AGERA-acidSEQ ID NO 48 P295Myr-KLAKKL-amideSEQ ID NO 49 P355Myr-LKLAKK-amideSEQ ID NO 50 P403Myr-LKLAK-[K]D-amideSEQ ID NO 51 P407Myr-AKKAL-[K]D-amideSEQ ID NO 52 P415Myr-KKKIII-amideSEQ ID NO 53 P397Pal-AARK-amideSEQ ID NO 54 P333Pal-KKLAK-amideSEQ ID NO 55 P380Myr-KKALKL-amideSEQ ID NO 56 P245Myr-GHR-amide P250Pal-GHR-amide P303Myr-GRK-amide P366Pal-G-H-[R]D-amide P368Pal-KAKL-amideSEQ ID NO 57 P370Pal-LAKK-amideSEQ ID NO 58 P360Myr-[K]D-K-[A]D-L-[K]D-amideSEQ ID NO 59 P373Myr-KKSLK-amideSEQ ID NO 60 P376Myr-LAIKK-amideSEQ ID NO 61 P369Myr-KKALKK-amideSEQ ID NO 62 P379Myr-KKALKA-amideSEQ ID NO 63 P381Myr-LKKALK-amideSEQ ID NO 64 P416Myr-IIIKKK-amideSEQ ID NO 65 P383Myr-SDGR-amideSEQ ID NO 66 P289Myr-LKAK-amideSEQ ID NO 67 P247Pal-VGVAPG-amideSEQ ID NO 68 P288Myr-KAKA-amideSEQ ID NO 69 P287Myr-AKAK-amideSEQ ID NO 70 P306Pal-GRKG-amideSEQ ID NO 71 P294Myr-GRKG-amideSEQ ID NO 72 P292Myr-LAKK-amideSEQ ID NO 73 P249Pal-GQPR-amideSEQ ID NO 74 P226Myr-KLAKK-amideSEQ ID NO 75 P261Pal-KLAKK-acidSEQ ID NO 76 P225Myr-KKGEM-amideSEQ ID NO 77 P216Myr-KRGKP-amideSEQ ID NO 78 P260Pal-KRGDR-acidSEQ ID NO 79 P214Myr-KKALK-amideSEQ ID NO 80 P329Pal-KKALK-amideSEQ ID NO 81 P262Pal-KKALK-acidSEQ ID NO 82 P215Myr-KKLAK-amideSEQ ID NO 83 P264Pal-GRKGD-acidSEQ ID NO 84 P222Myr-GRKGD-amideSEQ ID NO 85 P234Myr-KLAKKL-acidSEQ ID NO 86 P350Myr-AKKLAK-amideSEQ ID NO 87 P251Myr-AKKALK-acidSEQ ID NO 88 P230Myr-STKTTK-amideSEQ ID NO 89 P239Myr-SRVSRRSR-amideSEQ ID NO 90 P187Myr-LAKLAKKAF-amideSEQ ID NO 91 P189Myr-LAKKALKAF-acidSEQ ID NO 92 P285Myr-d-[KLAKKL]-acidSEQ ID NO 93 P243Myr-TKTSKS-amideSEQ ID NO 94 P238Myr-KRGDR-amideSEQ ID NO 95 P241Myr-KSSKS-amideSEQ ID NO 96 P242Myr-KTTK-amideSEQ ID NO 97 P359Myr-KKAL-d-[K]-amideSEQ ID NO 98 P235Myr-LKKALK-acidSEQ ID NO 99 P290Myr-KAKL-amideSEQ ID NO 100 P392Pal-R-amide P386Myr-DD-acid P391Myr-R-amide P316Pal-KK-amide P318Pal-RR-amide P393Myr-RH-amide P300Myr-GR-amide P315Myr-KK-amide P320Pal-HR-amide P347Myr-KK-acid P367Pal-K-[K]D-amide P420Myr-III-amide
The amino acid sequences in OABs of the present invention can be made synthetically by techniques well-known to those of skill in the art, including solid state peptide synthesis as described, for example, in U.S. Pat. No. 6,620,419. (Granted US patents and published US patent applications referenced herein are, to the extent pertinent, incorporated by reference.) Unless otherwise noted, amino acids in the OABs are in L form. As will be appreciated by persons having ordinary skill in the art amino acids are commonly represented by one or three letter sequences:
AAlaAlanineDAspAspartic AcidEGluGlutamic AcidGGlyGlycineHHisHistidineIIleIsoleucineKLysLysineLLeuLeucineMMetMethionineRArgArgininePProProlineSSerSerineTThrThreonineVValValine